LC–MS/MS Effectively Quantifies AB-PINACA in Human Hair

“We have developed and validated a method for detection of AB-PINACA in hair using a liquid chromatographytandem mass spectrometry system and applied it to head and pubic hair obtained in a case of intoxication.”

The recreational use of psychoactive substances to achieve changes in perception and mood is increasing. Although alcohol and caffeine continue to be the agents most commonly used for this purpose, consumption of more potent psychoactive substances has increased dramatically in recent years as they have become more readily accessible, e.g., through the internet.

To avoid the risk of criminalization carried by the use of illegal psychoactive drugs, such as cannabis and cocaine, new psychoactive substances (NPS) are continually being created. These agents mimic the effects of illicit drugs but have slightly different chemical structures that mean they are not covered by a list of prohibited substances and avoid control. Although NPS can carry a serious risk of psychosis, seizures, and coma, they are freely sold with claims that they provide a legal and safe way to achieve stimulatory and hallucinogenic effects — ‘legal highs’. The exact chemical composition of synthetic psychoactive substances is constantly changing to keep ahead of the law. Consequently, being an unknown entity, they pose a health risk to those who consume them and challenges to those trying to detect them, such as forensic chemists and toxicologists.

Not only the identification and quantification of new street drugs themselves challenge law enforcement authorities, but also the metabolic pathway of new substances needs to be explored in order to prove their abuse. Many of the new psychoactive substances reaching the market recently have been synthetic cannabinoids. These are man-made compounds that mimic the effects of cannabis, or more specifically its main active constituent delta-9 tetrahydrocannabinol (THC).

The European Commission states, “[…] the chemical identification of many unknown substances found by customs and suspected to be NPS requires the use of more sophisticated analytical techniques such as NMR and high resolution MS”. Both methods are well-established and complement each other in the comprehensive characterization of new psychoactive substances and their metabolic pathways. Numerous scientific articles report successful structure elucidation by NMR and metabolite identification by Mass Spectrometry like in the case of the 7-azaindole-derived synthetic cannabinoid 5F-AB-P7AICA. Bruker’s unique and comprehensive solution offering for forensic analysis enables law enforcement authorities worldwide to effectively fight the widespread distribution of prohibited substances.

A synthetic cannabinoid that has become an increasingly popular recreational drug is AB‐PINACA (N‐[(2S)‐1‐Amino‐3‐methyl‐1‐oxobutan‐2‐yl]‐1‐pentyl‐1H‐indazole‐3‐carboxamide). AB-PINACA binds to human cannabinoid receptors up to fourteen times more strongly than THC, causing agitation, confusion, lethargy, tachycardia, and hypertension. It has no therapeutic use and so is only available from the clandestine manufacture of ‘legal highs’ and as a research chemical.

Although analytical methodologies have been validated for the detection of AB-PINACA, it is not always detectable in biological samples during conventional toxicological screening. It is thus desirable to be able to detect potential consumption of AB-PINACA more reliably.

Synthetic cannabinoids have been detected in human hair using liquid chromatography−tandem mass spectrometry (LC–MS/MS), but the methodology had not been used to screen for AB-PINACA. This has now been undertaken on samples obtained from the victim of a road traffic accident who had in his possession a bag containing a white powder and labeled “AB-PINACA, 10 g, legal research chemical reagent use only”. The subject was found confused, with dilated pupils and slow speech but tested negative for alcohol.

Analysis of the white powder by 1H nuclear magnetic resonance using a Bruker AVANCE 300 spectrometer confirmed that its chemical structure matched that reported for AB-PINACA and that there were no impurities present.

Blood, urine, and hair samples from the subject were analyzed using LC–MS/MS and the hair test validated for linearity, repeatability, reproducibility, and detection and quantitation limits according to the Societé Française deToxicologie Analytique guidelines.

AB‐PINACA was detected in both the blood and urine samples, but the level in urine was very low indicating that AB-PINACA is rapidly and totally metabolized. The concentration of AB‐PINACA was significantly higher in head hair than in pubic hair. This is the reverse of data published for other cannabinoids, which may be a reflection of reduced pubic hair contamination due to the low AB-PINACA content in urine. There is also the possibility of external contamination of the head hair from hands that have touched AB-PINACA or AB-PINACA smoke in the environment.

The validation of a sensitive method for the detection of AB-PINACA in human hair provides the potential to determine the current or historical use of AB-PINACA with a minimally invasive procedure.

Arbouche N, et al. Testing for AB-PINACA in human hair: Distribution in head hair versus pubic hair. Drug Test Anal. 2019;11:610–616. https://onlinelibrary.wiley.com/doi/abs/10.1002/dta.2564